PROJECT SUMMARY/ABSTRACT Zika virus (ZIKV) is a mosquito transmitted flavivirus whose recent spread through the Americas has garnered worldwide concern; the WHO has declared Zika a Public Health Emergency of International Concern (1). Transmitted primarily by Aedes aegypti, infection with the 11 kilobase positive strand RNA virus can lead to mild, dengue like symptoms: fever and rash sometimes associated with conjunctivitis, arthralgia or myalgia. Recent outbreaks in French Polynesia and Central and South America have uncovered associations between Zika infection and neurological conditions (Guillain-Barre syndrome, (1, 2)) or birth defects (particularly miscarriage and microcephaly (1, 3)). With 440,000 to 1,300,000 suspected cases of ZIKV infection in Brazil in 2015 alone and 26 countries and territories in the Americas now reporting active Zika transmission, it is clear that this epidemic will affect millions (4, 5). The explosive spread of Zika virus in the Americas and the association with neurological and birth defects require that a safe and effective Zika vaccine is sorely needed. The recent development of vector independent RNA based delivery systems represents a practical platform for rapid and inexpensive development of new vaccine candidates. RNA based antigen expression candidates may be generated and purified using a common fully synthetic process, which alleviates the need for time consuming antigen specific process development. Central to the utility of this platform is the development of an effective RNA delivery formulation; we have developed a nano-liposomal alum formulation that can enhance antigen expression following delivery of RNA by intramuscular injection in vivo. This technology represents a Rapid Response RNA Vaccine platform that can enable the facile generation of effective vaccines in response to new emerging infectious disease threats. This R21 proposal will employ our Rapid Response RNA vaccine platform for ZIKV vaccine development. Initial experiments will focus on generation of attenuated ZIKV RNA vaccine candidates and verification of viral protein expression in vitro. Constructs shown to generate viral proteins will be advanced into immunogenicity studies in mice, wherein vaccine RNA will be combined with a delivery formulation. Comprehensive analysis of the immune response, with priority given to candidates that generate ZIKV neutralizing antibody responses, will be used to select lead vaccine candidates. Lead ZIKV vaccines will be advanced to efficacy testing in an AG 129 murine lethal challenge model.